Motile cilia are present in the epithelial cells of brain ventricles, trachea, oviduct, and sperm flagella. They play important roles in vivo including maintaining cerebrospinal fluid (CSF) circulation, clearing particles and pathogens from the respiratory tract, and sperm motility. Defects in motile cilia result in multiple diseases such as hydrocephalus, situs inversus, male infertility and et al. Most motile cilia have a ?9+2? axoneme. The shared structure of motile cilia in different tissues suggests that common regulatory mechanisms exist to govern expression of cilia/flagellar components. We hypothesize that expression of the genes essential for motile cilia structure/function are regulated by similar mechanisms so that the proteins are assembled into the motile cilia in a coordinated and timely manner. Sperm flagella are special motile cilia. Besides the core axoneme, other accessory structures are also assembled during spermiogenesis, the process of spermatid differentiation. During this phase, a number of genes essential for sperm flagella assembly are up-regulated. However, the transcriptional regulation of this suite of genes is poorly understood. By analyzing putative transcription factor binding sites in the promoter regions of genes that are essential for sperm flagella structure/function, we identified SOX5 as a putative key controller of cilia/flagellar genes. Sox-5 is a member of a family of genes that shows homology to the high motility group (HMG) box region of the testis determining gene, SRY. The mouse Sox5 gene encodes two major transcripts: the longer Sox5 isoform, originally named L-Sox5, but most authors refer it as Sox5; and a shorter isoform, S-Sox5. The first exon of the S-Sox5 transcript is a non-translated exon;, and it is not present in the L-Sox5 transcript. The translated 48 kDa S-SOX5 protein lacks the N-terminal half of the 84 kDa L-SOX5 protein. The two SOX5 proteins have different tissue distributions. L-SOX5 is expressed in multiple tissues, including the cartilage, heart, brain, kidney, lung, and skeletal muscle, but not in the testis, and it plays important roles in regulating processes of embryonic development and cell fate determination. S-SOX5 protein is only expressed in tissues with motile cilia, including the brain, lungs, and particularly in the testis. In the testis, it is localized in the nuclei of post-meiotic round spermatids. It has been shown that S-SOX5 regulates expression of several genes highly expressed in the testis. We hypothesize that S-SOX5 regulates expression of a suite of genes that are essential for sperm flagella formation and function. To test this hypothesis, we propose the following Specific Aims:1. To establish the critical the role of S-SOX5 in flagellogenesis and male fertility; and 2. To establish the target genes regulated by S-SOX5 in male germ cells. The proposed studies will help clarify the combinatorial transcription factor code that governs the differentiation and structural transformation of mammalian male germ cells. The tools that will be developed in pursuit of the proposed research will allow us to expand the project to a RO1 application to elucidate the transcriptional mechanisms that control mammalian sperm maturation.